Infective endocarditis is a serious infection of the heart with mortality rates in excess of 20%. This disease is thought to occur when bacteria or other microbes gain access to the blood, attach to previously damaged heart valves, and multiply. The oral or viridans streptococci are a leading cause of this illness, and among this group, Streptococcus sanguinis is especially important. Prevention of endocarditis relies primarily on antibiotic prophylaxis for at-risk patients prior to dental procedures that are likelyto result in bacteremia. However, it is now recognized that most cases of endocarditis result from daily activities that lead to bacteremia, for which dental antibiotic prophylaxis provides no protection. The identification of virulence determinants could provide new targets for novel prophylactic measures capable of providing continuous protection for persons at risk without selecting for antibiotic resistance in the mouth. Toward this goal, we have performed multiple screens of S. sanguinis mutants for loss of endocarditis virulence in an animal model. These studies have identified a lipoprotein, SsaB, that is uniquely important for endocarditis virulence and is also a promising target for therapy or prevention. SsaB, like other members of the family to which it belongs, functions in Mn (and perhaps Fe) uptake, O2 tolerance, and virulence. There are a number of mechanisms by which Mn could contribute to O2 tolerance and virulence, but it is not clear exactly which mechanisms are employed or are most important in streptococci. We have made a number of recent advances in relation to SsaB's potential functions that are novel not just for S. sanguinis, but for any Streptococcus, including characterization of an Mn-dependent ribonucleotide reductase that is essential for aerobic growth, characterization of the biological relevance of superoxide dismutase activity remaining in an ssaB mutant, identifying physiologically relevant in vitro growth conditions that mimic findings from the rabbit endocarditis model, the observation of metal-dependent processing of the SsaB protein, and the finding of an important role for SsaB in acid tolerance. We propose to expand upon these novel findings by examining SsaB regulation and function. This will provide the first test in any Streptococcus of multiple hypotheses generated by our recent findings that are related to metal transport. Given that we know that the primary defect of an ssaB mutant is metabolic, an important part of our study will be an unprecedented metabolomic analysis of O 2 and acid-stressed cells grown under carefully controlled conditions. This will be followed by testing of selected strains in in vitro and in vivo models of oral colonization and endocarditis virulence to test the biological relevance of our findings. The results of this study promise to be directly applicable to a number of other important pathogens and oral colonizers that are known to require Mn. Moreover, this study promises to identify novel approaches to control oral health and prevent infective endocarditis.